Identification of the putative urinary intraspecific recognition pheromone of the caracal caracal caracal

Hailemichael Goitom, Aron (2017-03)

Thesis (PhD)--Stellenbosch University, 2017.

Thesis

ENGLISH ABSTRACT: Sheep husbandry makes a considerable contribution to the economy of South Africa. However, it has been pressure during the last few decades due to the rapidly growing numbers of predators such as the caracal, Caracal caracal, and the red jackal, Canis mesomelas. Currently, sheep farming is hardly viable in the arid southern parts of the country. When experimenting with various methods of controlling caracal numbers, sheep farmers found that this problem animal could be effectively lured into traps using the urine of another male or female caracal. The main disadvantage here is that a sheep farmer could incur serious stock losses before he is able to obtain a starting sample of urine from another source (e.g., another farmer). It was hypothesised that caracal urine contains a volatile organic substance, or substances (VOCs), that are involved in the semiochemical communication between members of this species. The objective of this investigation was to identify these putative attractants in the urine, for the subsequent formulation of a caracal lure composed of synthetic analogues of the natural VOCs. As sheep farmers reported that male and female urine were equally effective attractants, it was considered unlikely that the attracting agent could be a sex pheromone; rather, it could have an intraspecific signalling function. Efforts were made to identify all the VOCs present in caracal urine samples donated by farmers from different sheep farming areas of the country. A total of 191 VOCs, including five steroid hormones, were identified. The identification of 86% of these VOCs was corroborated by gas chromatographic-mass spectrometric (GC-MS) comparison of the natural substances with authentic synthetic analogues. Obligate proteinuria is known in certain animals, in which it is not an indicator of renal abnormality. In the house mouse, Mus domesticus, for example, so-called major urinary proteins (MUPs) bind dehydro-exo-brevicomin and 2-(sec-butyl)-4,5-dihydrothiazole elicit male aggression. The carboxylesterase-like urinary excreted pheromone-binding protein, also known as cauxin, has been identified in the urine of several felids, but was not found in caracal urine. However, the pheromone-binding proteins ‘Transgelin fragment (M3WJ37)’ and ‘Uncharacterized protein (M3XEJ0)’, both previously identified in Felis catus and Felis silvestris catus, were identified in caracal urine. The macrocyclic C15 ketone, cyclopentadecanone, was identified as one of the VOCs present in the headspace gas of caracal urine. The headspace concentration of this ketone increased when the urine was heated at 95 ºC for 10 min. A similar, but more pronounced increase in cyclopentadecanone concentration was observed when urinary protein present in caracal urine was denatured by subjecting it to similar treatment. This was interpreted as an indication that this ketone is a ligand of caracal urinary protein. The C13, C14, C16, and C17 macrocyclic ketones were subsequently also identified as urinary protein ligands. In bioassays, a mixture of synthetic analogues of a large number of the other ketones identified in caracal urine, including cyclopentadecanone, elicited typical feline behaviour in two male caracal in captivity. Similar behavioural patterns were observed when a mixture of the C13, C14, C15, and C16 macrocyclic ketones were tested. It was concluded that these ketones, that are ligands of the caracal’s urinary protein, could play an essential role in the intraspecific communication of the caracal.

AFRIKAANSE OPSOMMING: Skaapboerdery maak ‘n aansienlike bydrae tot die Suid-Afrikaanse ekonomie, maar is die afgelope paar dekades onder toenemende druk as gevolg van die vinnige toename in rooikat- (Caracal caracal) en rooijakkals- (Canis mesomelas) bevolkings. Gevolglik is dié bedryf kwalik volhoubaar in die droë suidelike dele van die land. Skaapboere wat metodes ondersoek het om die skade wat rooikatte gereeld onder hulle skaapkuddes aanrig, het gevind dat hierdie probleemdier baie effektief gevang kan word deur die urien van ander wyfies of mannetjies as lokmiddel in vanghokke te gebruik. ‘n Nadeel van hierdie bestrydingsmetode is dat boere groot verliese kan ly voordat ‘n monster rooikat-urien bekom kan word om met so ‘n bestrydingsprogram te begin. Gebaseer op die hipotese dat rooikat-urien moontlik ‘n verbinding, of verbindings bevat wat ‘n rol in die semiochemiese kommunikasie van hierdie diere speel, is ‘n navorsingsprojek van stapel gestuur met die doel om sodanige verbinding(s) te identifiseer, ten einde ‘n lokstof uit sintetiese analoë te formuleer. Aangesien skaapboere gevind het dat die urien van wyfies en mannetjies ewe effektiewe lokmiddels is, is voorlopig aanvaar dat dit hier om intraspesifieke kommunikasie gaan en dat steroïdhormone, wat waarskynlik in die urien aanwesig is, dus waarskynlik nie die hoofrol in die lokwerking speel nie. Daar is gepoog om die chemiese samestelling van rooikat-urien so volledig as moontlik te bepaal deur al die vlugtige organiese verbindings (VOVs) in urien van verskillende geografiese gebiede waar met skape geboer word, te identifiseer. ‘n Totaal van 191 VOVs, wat die vyf steroïedhormone ingesluit het, is geïdentifiseer. Die identifikasie van 86% van hierdie verbindings is bevestig deur middel van gaschromatografies-massaspektrometriese vergelyking van die natuurlike verbindings met sintetiese analoë. Obligate proteinuria kom by sommige soogdiere voor, maar is nie ‘n indikator van renale nierversaking nie. In die huismuis, Mus domesticus, byvoorbeeld, bind die sogenaamde major urinêre proteïene (MUP) dehidro-exo-brevikomien en 2-(sek-butiel)-4,5-dihidrothiasool wat aggressie in manlike muise ontlok. The karboksielesterase-soortige urinêre feromoonbindende proteïen wat as ‘cauxin’ bekend staan, is in die urine van verskeie katspesies geïdentifiseer, maar is nie in rooikat-urien gevind nie. Die twee feromoonbindende proteïene ‘Transgelin fragment (M3WJ37)’ en ‘Uncharacterized protein (M3XEJ0)’, wat voorheen in Felis catus en Felis silvestris catus, geïdentifiseer is, is nou ook as feromoon-bindende proteïene in rooikat-urien gevind. Siklopentadekanoon is een van die VOVs wat in die bodamp van rooikat-urien gevind is. Met die verhitting van die urien by 95 °C vir 10 minute, het die bodampkonsentrasie van hierdie ketoon toegeneem. ‘n Soortgelyke, maar duideliker toename in bodampkonsentrasie is waargeneem wanneer die urinêre proteïen eers geïsolee is. Dit is as aanduiding aanvaar dat hierdie ketoon ‘n ligand van die rooikat se urinêre proteïen is. Die C13, C14, C16 en C17 makrosikliese ketone is later as verdere ligande van die proteïen geïdentifiseer. ‘n Mengsel van sintetiese analoë van verskeie ketone wat in rooikat-urien geïdentifiseer is en wat siklopentadekanoon ingesluit het, het die tipiese gedrag van kat-soortige diere ontlok in twee rooikatmannetjies wat in gevangenskap aangehou word. Soortgelyke gedragspatrone is weergeneem in toetse met ‘n mengsel van die C13, C14, C15 en C16 makrosikliese ketone. Daar is tot die gevolgtrekking gekom dat hierdie ketone, wat ligande van die rooikat se urinêre protein is, besmoontlik ‘n essensiële rol in die intraspesifieke chemiese kommunikasie van die rooikat speel.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/101159
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